Project Summary/Abstract Type2 diabetes (T2D) is the result of reduced insulin release from the pancreas following systemic insulin resistance. Insulin-producing ?-cells within the islets of Langerhans are responsible for maintaining blood glucose homeostasis; and dysfunctional insulin release in T2D results from reduced ?-cell function and mass. Insulin secretion from ?-cells is regulated by coordinated electrical signaling across the islet, which is established by Connexin-36 containing gap junctions (GJs). GJs ensure organized calcium signaling, resulting in proper insulin secretion and ?-cell survival. Preliminary data shows that GJs are prone to uncoupling upon exposure to a pro- inflammatory environment and in response to elevated circulating free fatty acids (FFAs). Given that T2D onset is associated with increases in both adipocyte-derived circulating pro-inflammatory cytokines and FFAs, we hypothesize that FFAs and pro-inflammatory cytokines induce islet dysfunction via FFA receptor (GPR40)-mediated GJ uncoupling, which ultimately induces altered Ca2+ signaling, reduced insulin secretion and ?-cell death. Furthermore, we hypothesize that preventing GJ uncoupling under these conditions can prevent diabetes-induced islet dysfunction. To test these hypotheses, both mouse and human islets will be exposed to concentrations of cytokines and palmitate that mimic the pre-diabetic environment. FFA signaling mechanisms will be delineated in islets treated with chemical antagonists/agonists of GPR40 and known GPR40 effector molecules (including phospholipase-C and protein kinase-C? pathways). We will also examine islets isolated from pre-diabetic, leptin receptor deficient mice (?db/db?) that model T2D. In an effort to prevent GJ uncoupling and associated islet dysfunction in T2D, we will synthesize novel inhibitory mimetic peptides targeted against Cx36 motifs that regulate coupling in response to FFAs/cytokines. In each case, state-of-the-art microscopy techniques will be used in conjunction with established methods to assess islet function by measuring GJ coupling, calcium signaling, insulin secretion and ?-cell viability. Results from these studies will define mechanisms by which islet dysfunction occurs in early in the disease that could be precipitating factors to T2D onset. We will also delineate opportunities for novel therapeutic interventions to preserve ?-cell mass, insulin secretory function and improve glycemic control in type2 diabetic patients.